间充质干细胞与神经干细胞序贯移植修复大鼠脊髓损伤的作用研究

发布时间：2018-01-27 10:24

本文关键词： 脊髓损伤细胞培养移植慢病毒转染序贯移植　出处：《安徽医科大学》2017年硕士论文　论文类型：学位论文

【摘要】：目的:通过比较由单独神经干细胞(NSCs)移植治疗大鼠脊髓损伤(SCI)和由骨髓间充质干细胞(BMSCs)与NSCs序贯治疗大鼠SCI后大鼠脊髓恢复情况的不同,研究BMSCs对大鼠脊髓微环境的影响以及序贯治疗的可行性和优势。方法:体外分离,培养,纯化NSCs和BMSCs,慢病毒转染标记神经干细胞,改良Allen法制造大鼠脊髓损伤(SCI)模型,将48只大鼠随机分为单独标记神经干细胞移植组(A组,n=24),间充质干细胞+标记神经干细胞序贯移植组(B组,n=24)。A组大鼠在T9脊髓损伤后3d经尾静脉注射单细胞浓度为/101ml6?的POLY-M慢病毒转染后的NSCs条件培养液2ml;B组大鼠在T9脊髓损伤后1d经尾静脉注射单细胞浓度为/101ml6?的BMSCs条件培养液1ml,脊髓损伤后3d经尾静脉注射单细胞浓度为/101ml6?的POLY-M慢病毒转染后的NSCs条件培养液2ml。每3只大鼠一组分笼饲养,由两名未参与分组的研究生对两组大鼠进行BBB评分,记录数据。分别于移植当天,移植后3天、1周、2周,3周、4周、6周、8周各取出A、B组3只大鼠,灌注法取脊髓损伤区的脊髓组织,福尔马林固定,制成石蜡切片,行HE染色和MAP-2以及GFAP免疫组化染色,观察脊髓空洞形成情况,神经干细胞分化生长情况,以及脊髓胶质疤痕大小。结果:序贯移植组BBB评分比单独神经干细胞移植组高(P0.05);观察大鼠在治疗后1,2,3,4,6,8周时的神经元细胞数目,血管生长情况,胶质疤痕以及脊髓空洞大小A,B组之间有显著性差异(P0.05)。结论:BMSCs可促进了NSCs的有效分化,减少胶质化倾向,改善了脊髓的微环境有利于治疗SCI,骨髓间充质干细胞与神经干细胞序贯移植对大鼠脊髓损伤的疗效优于单独移植神经干细胞。
[Abstract]:Objective: To compare the separate neural stem cells (NSCs) transplantation for the treatment of spinal cord injury (SCI) and bone marrow mesenchymal stem cells (BMSCs) and NSCs sequential treatment of rat SCI rat spinal cord after the recovery of the different effects of BMSCs on rat spinal cord microenvironment and the feasibility of sequential therapy and advantages. Methods: in vitro, separation, purification of NSCs and BMSCs, transfected with lentivirus labeled neural stem cells in rats with spinal cord injury, manufacturing method of modified Allen (SCI) model, 48 rats were randomly divided into separate labeled neural stem cells transplantation group (A group, n= 24), mesenchymal stem cells sequential + labeled neural stem cells transplantation group (group B, n=24).A T9 in rats after spinal cord injury 3D by intravenous injection of single cell concentration is /101ml6? POLY-M lentivirus transfected NSCs conditioned medium 2ml; rats in the B group at T9 after spinal cord injury 1D by caudal vein injection of single cell the concentration of /101ml6? The conditioned medium of BMSCs 1ml, 3D after spinal cord injury by intravenous injection of single cell concentration is /101ml6? POLY-M lentivirus transfected NSCs conditioned medium 2ml. 3 rats each group housed separately by two graduate students did not participate in the group of BBB scores of the two groups of rats were recorded data on the day of transplantation. After transplantation, 3 days, 1 weeks, 2 weeks, 3 weeks, 4 weeks, 6 weeks, 8 weeks each take A, 3 rats in group B, spinal cord, and spinal cord injury zone perfusion method in formalin fixed, paraffin sections were stained with HE and MAP-2 and GFAP immune immunohistochemical staining, syringomyelia formation, differentiation of neural stem cell growth, and spinal cord glial scar size. Results: sequential transplantation group BBB score than single neural stem cell transplantation group (P0.05); the number of cells in the neurons were observed after 1,2,3,4,6,8 weeks of treatment the rats, blood vessel growth, and glial scar The spinal cord cavity size A, there was significant difference between B groups (P0.05). Conclusion: BMSCs can promote the differentiation of NSCs effectively, reduce glial tendency, improve the micro environment of the spinal cord is conducive to the treatment of SCI, bone marrow mesenchymal stem cells and neural stem cells transplantation on the sequential curative effect is better than that of spinal cord injury rats injury transplantation of neural stem cells.

【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R651.2

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